Various organisms have at least one enzyme that degrades the RNA of RNA-DNA hybrids. These ribonucleases H (RNases H), so far, fall into two classes based upon primary amino acid sequence similarity. From our studies, we know that the well- characterized Escherichia coli RNase HI has homologs in many different species including human and mouse. We know that these mammalian proteins resemble in sequence and function the RNase H1 of Saccharomyces cerevisiae by having a double- stranded RNA-binding activity in addition to the RNase H activity. Synthesis of mRNA for RNase H1 of S. cerevisiae is constant throughout the cell cycle whereas the mRNA for RNase H2 is most abundant at S and G2/M phase. Double-deletion mutant strains are viable yet there are diffferences in sensitivities of the single mutant strains to caffeine and hydroxyurea. The level of mRNA and RNase H2(35) protein increase in the absence of RNase H1. These results suggest that RNase H1 is involved in housekeeping function, removing inadvertently formed RNA-DNA hybrids while RNase H2 is involved in DNA replication and repair. - Ribonuclease H, cell cycle, DNA replication, DNA repair